ABSTRACT Within tumor beds, T and B cells often interact to form highly organized structures similar to lymph nodes, termed tertiary lymphoid structures (TLS), which are associated with better outcomes in many tumors. TFH cells are crucial for the formation of germinal centers and humoral responses, and our new data show that TFH cells become the main producers of CXCL13 and TNFS14/LIGHT upon vaccination. The assembly and maintenance of TLS should be therefore dependent on TFH responses. Our new data demonstrate that Special AT-rich sequence-binding protein-1 (Satb1) ablation specifically in T cells leads to enhanced TFH differentiation and augmented Ag-specific humoral responses, which is associated with concurrent ICOS and PD-1 de-repression. Accordingly, our central hypothesis is that LIGHT+CXCL13+ TFH cell formation and, subsequently, the orchestration of TLS in cancer, is governed by Satb1 silencing in CD4 T cells by both de-repressing ICOS in TFH cells and suppressing Foxp3+PD-1highCXCR5+ T follicular regulatory (TFR) cell formation through PD-1 up-regulation. Therefore, TGF-? paradoxically enhances the generation of TFH cells and the formation of TLS through Satb1 repression. In Aim 1, we will define the role of SATB1-dependent ICOS expression during TFH differentiation. Through ChIP-PCR and functional analysis of Satb1-competent vs. Satb1- deficient T cells in vivo, we will substantiate a novel epigenetic mechanism whereby the master genomic organizer Satb1 governs ICOS expression, leading to enhanced TFH differentiation in the absence of Satb1. In Aim 2, we will determine the role of SATB1 in TGF-?-driven, Treg-dependent TFH differentiation. Here, we will combine geentic manipulation and existing transgenic models to establish to what extent the mechanism of TGF-?-driven TFH differentiation is Satb1- and PD-1- dependent, in a manner that requires decreased TFR formation. In Aim 3, we will recapitulate the mechanisms leading to the formation and protective activity of TLS in vivo in ovarian cancer. By leveraging unique transgenic models, our ovarian cancer- specific CAR T cells and our viable single-cell suspensions from freshly dissociated ovarian carcinomas, we will define a novel TGF-? ? Satb1 silencing ? TFH cell formation axis driving relevant anti-tumor humoral responses. Our work will exert a profound effect in the field by elucidating how epigenetic programs controlled by SATB1 govern the generation of TFH cells at tumor beds in a TGF-?-dependent manner. Recapitulating these mechanisms in vivo will pave the way for more effective immunotherapies aimed to promote combined humoral and T cell responses through the orchestration of TLS in irresectable/metastatic tumors, and could lead to the identification of antibodies with anti-tumor activity spontaneously produced at tumor beds.